Enediyne natural products are characterized by the presence of the enediyne ring structure also referred to as the warhead. The labile enediyne ring structure undergoes a thermodynamically favorable Bergman cyclization resulting in transient formation of a biradical species. The biradical species is capable of inducing irreversible DNA damage in the cell. This reactivity gives rise to potential biological activity against both bacterial and tumor cell lines. Enediynes have potential as anticancer agents because of their ability to cleave DNA. Calicheamicin is currently in clinical trials as an anticancer agent for acute myeloid leukemia (Nabhan C. and Tallman M S, Clin Lymphoma (2002) Mar;2 Suppl 1: S19-23). Enediynes also have utility as anti-infective agents. Accordingly, processes for improving production of existing enediynes or producing novel modified enediynes are of great interest to the pharmaceutical industry.
Enediynes are a structurally diverse group of compounds. Chromoprotein enediynes refer to enediynes associated with a protein conferring stability to the complex under physiological conditions. Non-chromoprotein enediynes refer to enediynes that require no additional stabilization factors. The structure of the chromoprotein enediynes neocarzinostatin and C-1027, and the non-chromoprotein enediynes calicheamicin and dynemicin are shown below with the dodecapolyene backbone forming the warhead structure in each enediyne highlighted in bold.

Efforts at discovering the genes responsible for synthesis of the warhead structure that characterizes enediynes have been unsuccessful. Genes encoding biosynthetic enzymes for the aryltetrasaccharide of calicheamicin, and for calicheamicin resistance are described in WO 00/37608. Additional genes involved in the biosynthesis of the chromoprotein enediyne C-1027 have been isolated (Liu, et al. Antimicrobial Agents and Chemotherapy, vol. 44, pp 382-292 (2000); WO 00/40596). Isotopic incorporation experiments have indicated that the enediyne backbones of esperamicin, dynemycin, and neocarzinostatin are acetate derived (Hansens, O. D. et al., J. Am. Chem Soc. 11, vol 111 pp. 3295-3299 (1989); Lam, K. et al., J. Am. Chem. Soc. vol. 115, pp 12340-12345 (1993); Tokiwa, Y et al., J. Am. Chem Soc. vol. 113 pp. 4107-4110). However, both PCR and DNA probes homologous to type I and type II PKSs have failed to identify the presence of PKS genes associated with biosynthesis of enediynes in known enediyne producing microorganisms (WO 00/40596; W. Liu & B. Shen, Antimicrobial Agents Chemotherapy, vol. 44 No. 2 pp. 382-392 (2000)).
Elucidation of the genes involved in biosynthesis of enediynes, particularly the warhead structure, would provide access to rational engineering of enediyne biosynthesis for novel drug leads and makes it possible to construct overproducing strains by de-regulating the biosynthetic machinery. Elucidation of PKS genes involved in the biosynthesis of enediynes would contribute to the field of combinatorial biosynthesis by expanding the repertoire of PKS genes available for making novel enediynes via combinatorial biosynthesis.
Existing screening methods for identifying enediyne-producing microbes are laborious, time-consuming and have not provided sufficient discrimination to date to detect organisms producing enediyne natural products at low levels. There is a need for improved tools to detect enediyne-producing organisms. There is also a need for tools capable of detecting organisms that produce enediynes at levels that are not detected by traditional culture tests.